Adherent cell requirement for the effect of vitamin E on in vitro antibody synthesis.

Mice deficient in dietary vitamin E are impaired in their humoral and cell-mediated immunological responses. The basis for this impaired immunocompetence was investigated by using the in vitro antibody response as an assay system. Spleen cells from mice fed vitamin E-deficient diets were low responders to the antigens, sheep red blood cells (SRBC) and dinitrophenyl-L-lysine-Ficoll (DNP-Ficoll). However, they responded as well as mice fed vitamin E-supplemented diets to the relatively macrophage-independent antigen trinitrophenylated-lipopolysaccharide (TNP-LPS). This suggested that the macrophage was the cell most affected by the vitamin E deficiency. The involvement of macrophages was confirmed directly by mixing experiments, in which it was shown that macrophages from vitamin E-deficient mice were unable to support an antibody response by macrophage-depleted spleen cells from vitamin E-supplemented mice. Macrophages from vitamin E-deficient mice expressed less Ia antigen, and seemed less able to present antigen to nonadherent cells. However, it was found that macrophages from vitamin E-deficient mice not only lacked accessory cell function, but could act instead as suppressor cells. The effect of dietary vitamin E was noted with either saturated or unsaturated sources of fat in the diet.

The effect of vitamin E deficiency on some platelet membrane properties.

The effects of alpha-tocopherol (vitamin E) deficiency on membrane properties of platelets were studied to determine if vitamin E has a measureable stabilizing role in biological membranes. Three groups of rats and three of mice were studied: two groups consisted of Fisher strain rats and one of Sprague-Dawley rats fed a Draper corn oil diet with and without high levels of supplementary vitamin E. The mice were two groups of BALB/c animals maintained on an 8% hydrogenated coconut oil diet, and one group of CBA/J mice on an 8% lard diet, in each case either deficient in or supplemented with vitamin E. The relative content of fatty acids obtained from both rat platelets and erythrocytes was unchanged by vitamin E deficiency. Depletion of vitamin E had no effect on the degree of fluorescence polarization of 1,6-diphenyl-1,3,5,-hexatriene-labeled rat platelets. No changes in hematocrit values were seen in any of the studies. The platelet count of only the vitamin E-deficient Sprague-Dawley rats was elevated with respect to vitamin E-supplemented counterparts; the others remained constant. Platelet reactivities, as measured by ADP-and thrombin-induced platelet aggregation and by the thrombin-induced changes in platelet transmembrane potential, were unaffected by vitamin E deficiency in all three groups of rats. Our results indicate that a membrane stabilizing effect of vitamin E on rat platelet or erythrocyte membrane fatty acids or on platelet response to external stimuli could not be demonstrated, nor was elevation in platelet count a general phenomenon associated with vitamin E deficiency.-Whitin, J. C., R. K. Gordon, L. M. Corwin, and E. R. Simons. The effect of vitamin E deficiency on some platelet membrane properties.

Vitamin E protection against tumor formation by transplanted murine sarcoma cells.

Mice fed vitamin E at a level of 0.5 g DL-alpha-tocopheryl acetate/kg diet demonstrated decreased incidence and rate of appearance of tumors produced by transplanted sarcoma cells (K3T3), compared to control groups fed diets without the vitamin supplement. Protection was dependent on the degree of unsaturation of dietary fat and on the size of the tumor cell challenge. When vitamin E was increased 10-fold (to 5 g/kg diet), the protective effect was no longer observed. Protection may be mediated through the host immune system, because sublethal, whole-body X-irradiation abrogated differences in tumor development between the +E and the -E mice. Studies with in vitro immunization showed that treatment of the K3T3 cell with vitamin E enhanced its ability to induce a cytotoxic response. It appears that the direct effect of vitamin E is on the tumor cell rather than on the immune system, since spleen cells from mice fed diets with and without vitamin E supplementation were indistinguishable in their response to untreated K3T3 cells. K3T3 cells treated with excessive levels of vitamin E were unable to induce a cytotoxic response, a result that correlates with the loss of protection against tumor development when massive doses of vitamin E were fed.

Influence of vitamin E on the mitogenic response of murine lymphoid cells.

The effect of vitamin E on mitogenesis by polyclonal activators was studied and the vitamin was found to be stimulatory but selective in its action. Vitamin E itself is a mitogen for murine spleen cells. At suboptimal vitamin concentrations, it was capable of stimulating the response to low levels of the thymus-dependent lymphocyte (T cell) mitogen, concanavalin A (conA), but not when conA was itself at optimal levels. When vitamin E was added to the diet at normal levels, it was not as effective in stimulating mitogenesis as it was at much higher levels. The effect of the vitamin on T cell mitogenesis could be modified by the degree of unsaturation of the dietary fat; it was more effective when dietary polyunsaturated fatty acids (PUFA) were low. Under several conditions, it was shown that vitamin E can increase the phytohemagglutinin (PHA)/conA response ratio, which may suggest an effect of the vitamin on the maturation of T cells. In normal mice, vitamin E also stimulated the response to lipopolysaccharide (LPS), a "bursa-equivalent" lymphocyte (B cell) mitogen, but it was unable to do so when spleen cells from athymic, nude mice were used. This suggests a requirement for thymic factors in order for vitamin E to stimulate mitogenesis of B cells.

Effect of L-azetidine 2-carboxylic acid on growth and proline metabolism in Escherichia coli.

The effects of L-azetidine 2-carboxylic acid on growth and proline metabolism in a proline-requiring auxotroph of Escherichia coli are described. The homologue inhibited growth of the wild type and it, alone, did not substitute effectively for proline as a growth supplement for the mutant. In medium containing 0.05 mM proline, the addition of increasing amounts of homologue progressively inhibited growth of the wild type but stimulated growth of the mutant at homologue: proline ratios of 10 : 1 and 50 : 1. This suggested that the homologue exerted a "sparing effect" on proline in the mutant. The incorporation of L-[U-14C]proline and L-[3H]azetidine 2-carboxylic acid into hot trichloroacetic acid-insoluble material in the mutant was measured. Amino acid analysis of the insoluble material from cells incubated with radiolabeled proline alone revealed that proline was partially degraded and metabolized to other amino acids prior to incorporation into protein. The addition of unlabeled homologue to the incubation medium significantly reduced proline catabolism, suggesting that the homologue exerted a sparing effect on proline in this mutant. In medium containing unlabeled proline and radiolabeled L-azetidine 2-carboxylic acid, the homologue was incorporated both intact and partially degraded prior to incorporation into protein. Alanine was the major L-azetidine 2-carboxylic acid catabolite.

Mutations affecting aromatic amino acid transport in Escherichia coli and Salmonella typhimurium.

A genetic locus, aroT, located between chr and the trp operon in Salmonella typhimurium, and similar genes, aroR and aroS, near the trp locus of Escherichia coli, were found to be involved in the transport of aromatic amino acids. Genetic lesions at these loci cause a variable diminution in uptake and accumulation of aromatic amino acids, alanine and glycine compared with the wild type. The F'trp episome carries the aro R locus. Curing an E. coli strain of the F'trp episome which covers a chromosomal deletion from cysB through the trp operon and tonB regions, results in a 60 to 80% decrease in tryptophan uptake. The introduction of F'trp into a trp operon-deleted S. typhimurium of low transport ability restores transportability, suggesting that aroT in this organism may be homologous with aroR in E. coli. In E. coli, tryptophan accumulation is normally increased by prior growth in L-tryptophan, while in S. typhimurium it is repressed. In both genera, the trpR gene appears to have no effect on the tryptophan transport capabilities in response to changes in the concentration of L-tryptophan in the medium. Tryptophan transport in the S. typhimurium F'trp hybrid was subject to repression, while in the E. coli strain which carries F'trp covering the equivalent chromosomal delection, an increase in tryptophan accumulation was shown after growth in L-tryptophan supplemented medium.